The long term goal of this proposal is to elucidate the molecular mechanisms governing the tissue specific gene expression during development and differentiation of ocular lens. The primary focus is to understand the regulation of gamma-crystallin gene expression which will be subsequently extended to the other crystallin genes. The proposed study will be pursued in the following manner: The gamma-crystallin protein(s) will be isolated from Xenopus lens and polyclonal antibodies will be prepared. These antibodies will be employed for the proposed studies. The genomic clones containing the gamma 1 crystallin gene will be subcloned into a plasmid vector and characterized by DNA sequence analysis, gene copy number and hybrid-select translation. The DNA region(s) essential for the tissue specific expression will be identified by preparing a series of deleted, mutated and insertion genes. These will be introduced into either homologous or heterologous lens cells to assay for the specific expression. The developmental expression of gamma-crystallin gene will be undertaken both at the RNA and protein level and the results will be compared with the morphological changes associated with lens development. A tissue culture system will be developed from Xenopus eye lens epithelial cells to provide a homologous cell culture system which will subsequently be used for the transfection assay.